An in vitro transcription system which faithfully reflects in vivo transcription has been developed. This proposal outlines a series of experiments which principally utilizes this system to study the production of adenovirus with primary transcripts and the processing and modification of those transcripts into mRNA. Briefly, these experiments involve in vitro transcription of Ad 2 DNA for short periods of time, followed by analysis of the RNA on polyacrylamide gels containing 98 percent formamide with specific Ad 2 DNA fragments. Other experiments involve the use of the specific DNA fragments in exhaustive hybridization studies which would serve to identify promote and termination sequences on Ad 2 DNA as well as provide information about the metabolism of various regulatory classes of Ad 2 RNA. These experiments should provide some understanding of the molecular events involved in the regulation of gene expression and provide a basis for the identification of some of the regulatory proteins involved in these processes. It is hoped that the elucidation of some of these regulatory controls will contribute to our understanding of cellular differentiation in general, and of cellular transformation due to DNA oncogenic viruses in particular. In addition, these studies should also contribute to our knowledge of the molecular pathogenesis of various diseases caused by viral interference of host cell metabolism.